Clinical studies of multiple pain disorders indicate that a patient's biological sex is a common risk factor for pain, with many studies demonstrating that women are at increased risk for pain in many acute and chronic conditions. The proposed translational clinical study directly tests the hypotheses that estradiol (172-E2) increases peripheral nociceptor activity in human acute pain patients and that polymorphisms of the estrogen receptors are associated with increased nociceptor activity and post-operative pain. To address this hypothesis, we have developed a method for evaluating estradiol enhancement of the exocytotic activity of peripheral human nociceptors innervating surgical biopsies of healthy tissue (evaluating substance P (SP) release from neuron terminals innervating dental pulp from extracted third molar teeth). Our preliminary studies identified estrogen receptor 1 (ER1) and ER1 in both human trigeminal ganglia neurons and in peripheral neurons in surgical biopsies, and discovered that estradiol significantly increases bradykinin/prostaglandin E2 (BK/PGE2)-evoked iSP from human tissue biopsies collected from women, but not from men. Moreover, in this A1 revision, we report new findings that certain polymorphisms of ER1 and ER2 are positively correlated with increased intensity of post-operative pain in these patients. Thus, we have developed a working hypothesis that estrogen sensitizes peripheral terminals of human trigeminal nociceptors. To evaluate this hypothesis, we propose the following aims: Specific Aim 1: Determine whether females differ from males for increased release of SP following stimulation with either BK/PGE2 or with capsaicin. Specific Aim 2: Determine whether elevated circulating estradiol levels are associated with increased release of SP following stimulation with either BK/PGE2 or with capsaicin Specific Aim 3: Determine whether specific polymorphisms of the estrogen receptor alpha (ER1) or beta (ER2) are associated with increased BK/PGE2- or capsaicin-evoked release of SP from surgical biopsies exposed to estradiol. Specific Aim 4: Establish a data and gene bank to facilitate future genetic epidemiological and pharmacogenetic studies. This database will be used for three proposed secondary analyses including an evaluation of the association between ER polymorphisms and post-surgical pain and will be also available to other investigators to foster the NIH policy for data sharing. These integrated studies will evaluate estrogen modulation of peripheral human nociceptors at the pharmacological, biochemical and genetic levels. Collectively, these studies provide a comprehensive evaluation of the hypothesis that activation of estrogen receptors enhances nociceptor function in women. [unreadable] [unreadable] PUBLIC HEALTH RELEVANCE: Although women are at greater risk for pain in many acute and chronic pain conditions, comparatively little is known about the mechanisms mediating this effect. In this study, we will test whether estrogens enhance pain nerve activity from surgical biopsies collected from female versus male patients and study whether certain natural mutations of the gene for the estrogen receptors are correlated with increased levels of post- operative pain. Understanding estrogen activity on pain nerves will not only increase our understanding of why women are at risk for pain, but offers the potential for developing new types of selective estrogen receptor modulators capable of reducing this effect. [unreadable] [unreadable] [unreadable]